Abstract
PAX3-FKHR is an oncogenic form of the developmental regulator Pax3 transcription factor. PAX3-FKHR results from a t(2,13) chromosomal translocation, a unique genetic marker of alveolar rhabdomyosarcoma. In this study, we showed that ectopic expression of PAX3-FKHR, but not Pax3, in fibroblasts altered cell cycle control and accelerated G(0)/G(1) to S cell cycle transition. PAX3-FKHR-expressing cells had reduced expression of p27(Kip1) protein, a key cell cycle regulator. The reduction in p27(Kip1) levels by PAX3-FKHR resulted from destabilization of p27(Kip1) as shown by cycloheximide treatment and in vivo pulse-chase labeling experiments. The reduced p27(Kip1) protein level in PAX3-FKHR-expressing cells was restored to the level of control cells by treatment with chemical inhibitors that specifically blocked 26 S proteasome activity. Along with the reduction in p27(Kip1) protein, PAX3-FKHR-expressing cells exhibited elevated expression of F-box Skp2 protein, a substrate-specific component of SCF (Skp1-Cullin-F box protein) ligase involved in the cell cycle-dependent control of p27(Kip1) ubiquitination and 26 S proteasome dependent degradation. Finally, we showed that ectopic expression of p27(Kip1) in PAX3-FKHR-expressing cells significantly reduced the proliferation and colony-forming potential of these cells, implicating that down-regulation of p27(Kip1) protein played an active role in the PAX3-FKHR-directed cell transformation.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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3T3 Cells
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Adolescent
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Animals
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CDC2-CDC28 Kinases*
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Cell Cycle / physiology
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / metabolism*
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Cell Transformation, Neoplastic*
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Child
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Cyclin-Dependent Kinase 2
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Cyclin-Dependent Kinase Inhibitor p27
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Cyclin-Dependent Kinases / metabolism
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism*
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Forkhead Box Protein O1
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Forkhead Transcription Factors
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Genes, Tumor Suppressor
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Humans
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Mice
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PAX3 Transcription Factor
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Paired Box Transcription Factors
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Peptide Hydrolases / metabolism*
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Proteasome Endopeptidase Complex*
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Protein Serine-Threonine Kinases / metabolism
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism*
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Rhabdomyosarcoma / metabolism
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S-Phase Kinase-Associated Proteins
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Transcription Factors / genetics
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Transcription Factors / metabolism*
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Tumor Suppressor Proteins / genetics
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Tumor Suppressor Proteins / metabolism*
Substances
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Cdkn1b protein, mouse
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Cell Cycle Proteins
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DNA-Binding Proteins
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FOXO1 protein, human
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Forkhead Box Protein O1
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Forkhead Transcription Factors
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Foxo1 protein, mouse
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PAX3 Transcription Factor
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PAX3 protein, human
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Paired Box Transcription Factors
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Recombinant Fusion Proteins
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S-Phase Kinase-Associated Proteins
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Transcription Factors
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Tumor Suppressor Proteins
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Pax3 protein, mouse
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Cyclin-Dependent Kinase Inhibitor p27
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Protein Serine-Threonine Kinases
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CDC2-CDC28 Kinases
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CDK2 protein, human
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Cdk2 protein, mouse
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Cyclin-Dependent Kinase 2
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Cyclin-Dependent Kinases
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Peptide Hydrolases
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Proteasome Endopeptidase Complex
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ATP dependent 26S protease